Chlamydia trachomatis is an obligate intracellular bacterial pathogen that is the cause of a wide spectrum of human diseases, including blinding trachoma. Chlamydiae infect mammalian cells by attachment, endocytosis and inhibition of lysosomal fusion with endosomes containing chlamydia. The target host cell in vivo is typically the columnar epithelial cell; however, little is known about the molecular mechanism of chlamydial infection of host cells. The long-term objective is to understand chlamydial pathogenesis and virulence in the context of the interaction of chlamydiae with their host cells. This will yield important fundamental information for (a) understanding the mechanism of infection, (b) mediators of virulence, and (c) the development of new approaches for intervention. The specific aims of this project will define molecular and biochemical mechanisms involved in attachment and invasion of chlamydia with host cells. The aims are derived from new information that demonstrates a novel and essential role for glycosaminoglycan-mediated attachment of chlamydia to host cells. The hypothesis is that chlamydiae attach to mammalian host cells by a trimolecular complex in which heparan sulfate-like glycosaminoglycan bridges lectin-like receptors on chlamydia and on the host cell. A molecular understanding of attachment will provide a rational foundation for the study of other important microbiological characteristics such as uptake, inhibition of lysosomal fusion, and persistence of infection. The immediate goals of our collaborative efforts are isolation and characterization of the glycosaminoglycan ligand. The purified glycosaminoglycan will be digested enzymatically and chemically, and analyzed by various "oligosaccharide mapping" techniques. Based on preliminary studies with model sulfated oligosaccharides, we anticipate that electrospray ionization mass spectrometry and tandem mass spectrometry will be extremely useful for molecular weight measurements and sequencing of these chlamydial samples.